Negative or positive cooperation in calcium binding to detergent-solubilized ATPase of the sarcoplasmic reticulum: Its modulation by a high concentration of ATP

Two different conformations of chemically equivalent Ca²⁺-ATPase molecules in the sarcoplasmic reticulum have been shown to non- and positive cooperatively bind two calcium ions, respectively (Nakamura, J. (1994) J. Biol. Chem. 269, 30822-30827). At pH 7.40, these ATP-ase molecules split into E₁ (high affinity state for calcium) and E₂ (low affinity state for calcium), respectively, before calcium binding. At this pH, calcium binding to the monomeric ATPase, solubilized with dodecyloctaethylenglycol monoether, was studied by examining ⁴⁵Ca²⁺ binding to the ATPase and calcium dependences of its phosphorylation, fluorescence intensity, ATP-hydrolysis at a low (5 μM) concentration of ATP, and acetyl phosphate hydrolysis. The results suggest that the solubilized ATPase molecules predominantly preexist in E₂ and negative cooperatively (the Hill value (n_H) = 0.5-0.6) bind 2 mol of calcium/mol of the ATPase with an apparent calcium affinity (K_0.5) of 3-5 μM. The nonequivalences of calcium bindings at the membranous ATPase molecules seem to result from the intermolecular interaction of the molecules. A high concentration (5 mM) of ATP modulated the binding manner so that it became positively cooperative (n_H ∼ 2) and increased the K_0.5 to 0.1 μM.